The pattern of RNA integrity and the expression of housekeeping genes are influenced by sodium hypochlorite and ascorbic acid.

BACKGROUND: Basic biological science research deals with nucleic acid isolation. Post-isolation nucleic acid integrity has a pivotal role in further elucidating gene expression and other molecular mechanisms. RNA (ribonucleic acid), cDNA (complementary deoxyribonucleic acid), and PCR (Polymerase chain reaction) products' integrity and quality are affected by several factors in biochemical and biophysical degradation modes. Inadequate evidence was noted about the direct effects of sodium hypochlorite and L-ascorbic acid. OBJECTIVES: This study aims to test the effects of sodium hypochlorite (SHC) and L-ascorbic acid (LAA) in total RNA and PCR products, respectively, in an acellular condition. METHODS: The study was categorized into three steps total RNA, cDNA, and PCR product evaluations. mBM-MSCs were used to extract RNA and then treated with SHC. Crude total RNA and, after DNase 1 treatment, the bands of total RNA samples were visualized by agarose gel electrophoresis. cDNAs were synthesized from SHC-treated (0.25%) and untreated RNAs, which were also expressed on the gel. LAA (5 µM, 15 µM, 25 µM, and 50 µM) were added to cDNAs synthesized from SHC- and non-SHC-treated samples. Housekeeping genes, Gapdh (Glyceraldehyde 3-phosphate dehydrogenase), and 18S rRNA (18S Ribosomal ribonucleic acid) were amplified in both groups. RESULTS: SHC-treated samples produced clearer bands on an agarose gel. Its treatment did not affect the integrated densities of agarose bands which revealed non-significant (P ≤ 0.05) differences in SHC-treated, untreated RNA, and cDNA. However, significant variations were observed at the PCR level. SHC-treated samples expressed decreased housekeeping gene expression in amplified products (Gapdh and 18S rRNA) and slightly but non-significantly high band intensities appeared in the presence of LAA. Significant variable differences (*P ≤ 0.05) were observed between SHC-treated and non-treated groups after LAA treatment. CONCLUSIONS: SHC (0.25%) is favorable in removing RNases and maintaining the integrity of RNA. cDNA synthesis did not affect by SHC treatment, and it follows the same as untreated samples after DNase 1 treatment. LAA drew a positive impact to improve the quality of PCR products in terms of band intensities, which is insignificant in SHC-treated RNA. Interestingly, it was revealed from our study that 5-25 µM LAA has the most beneficial role in the acquisition of PCR products, i.e. gene expression. These concentrations can be safely used to improve the quality of gene expression. This phenomenon can be used to achieve other, rarer, desired gene expressions. Further research is needed to explore the effects of SHC on the acquisition of PCR products using other solutions.

Angiogenic potential and wound healing efficacy of chitosan derived hydrogels at varied concentrations of APTES in chick and mouse models.

Chitosan (Cs) based biomaterials seem to be indispensable for neovasculogenesis and angiogenesis that ensure accelerated wound healing. Cs/poly (vinyl alcohol) (PVA) bio-constructs were cross-linked and investigated with varying concentrations of aminopropyltriethoxysilane (APTES). This study comprised of three phases: fabrication of hydrogels, characterization, assessment of angiogenic potential along with toxico-pathological effects, wound healing efficacy in chick and mice, respectively. The hydrogels were characterized by FTIR, SEM and TGA and the swelling response was examined in different solvents. The hydrogels swelling ratio was decreased with increasing amount of APTES, showed the highest swelling at acidic and basic pH while low swelling at neutral pH. Chorioallantoic membranes (CAM) assay was performed to study in-vivo angiogenesis, toxicological, morphological, biochemical and histological analyses in developing chicks. The results showed remarkably improved angiogenesis with little deviations in morphological, histological features and liver enzymes of chick embryos at higher concentrations of APTES. Besides, full thickness wounds were excised on mice dorsolateral skin to assess the wound healing. The rate of wound size reduction was significantly higher after topical application of hydrogels with elevated levels of crosslinker. Hence, the hydrogels showed enhanced angiogenesis, accelerated wound healing with little or no observable in-vivo toxicity.

Development and characterization of chitosan and acrylic acid-based novel biodegradable polymeric films for soil conditioning.

In this study, biodegradable polymeric films (BPFs) based on chitosan and acrylic acid cross-linked with 3-aminopropyl triethoxysilane (APTES) were developed for water retention and soil-conditioning applications in areas sufferings from water scarcity. A series of BPFs were prepared by varying the amount of silica nanoparticles (SiNPs) (0.67% to 2.6%) and a correlation of the optimum amount of SiNPs with thermal stability, morphology, swellability (at various pH), degradability, and anti-microbial activity were deduced. The obtained results showed that the NP 8 (containing 2.51% of SiNPs) exhibited the maximum absorption capacity (1815%) in distilled water, whereas NP6 (including 1.88% of SiNPs) expressed the maximum thermal stability (T50% at 375.61 °C). The microscopic images further strengthen this observation because the maximum number of micro-porous cavities was shown on the surface of NP8. The time-dependent swelling response in distilled water accomplished that hydrophilicity (percentage swelling) of films was enhanced with an increase in the concentration of SiNPs. All BPFs samples exhibited inhibitory response against both gram-positive (for Staphylococcus aureus was 2.9 cm for NP6) and gram-negative (for Escherichia coli was 0.9 cm for NP8) bacteria. The biodegradation test inferred that the degradation of BPFs in soil did not affect the soil fertility as nano-silica is proven as growth-promoting miniatures. It can be concluded that these BPFs may be efficiently employed in the agriculture sector for water retention and as a soil conditioner.

Inflammation targeted chitosan-based hydrogel for controlled release of diclofenac sodium.

Inflammation is a key challenge in the treatment of chronic diseases. Spurred by topical advancement in polymer chemistry and drug delivery, hydrogels that release a drug in temporal, spatial and dosage controlled fashion have been trendy. This research focused on the fabrication of hydrogels with controlled drug release properties to control inflammation. Chitosan and polyvinyl pyrrolidone were used as base polymers and crosslinked with epichlorohydrin to form hydrogel films by solution casting technique. Prepared hydrogels were analyzed by swelling analysis in deionized water, buffer and electrolyte solutions and gel fraction. Functional groups confirmation and development of new covalent and hydrogen bonds, thermal stability (28.49%) and crystallinity were evaluated by FTIR, TGA and WAXRD, respectively. Rheological properties including gel strength and yield stress, elasticity (2309 MPa), porosity (75%) and hydrophilicity (73°) of prepared hydrogels were also evaluated. In vitro studies confirmed that prepared hydrogels have good biodegradability, excellent antimicrobial property and admirable cytotoxicity. Drug release profile (87.56% in 130 min) along with the drug encapsulation efficiency (84%) of prepared hydrogels was also studied. These results paved the path towards the development of hydrogels that can release the drugs with desired temporal patterns.

Prevalence and association of HBV and HCV infection with cardiovascular disease risk factors in a peri-urban population.

OBJECTIVE: To investigate the possible associations of hepatitis B and C virus infection with cardiovascular disease risk factors inperi-urban population. METHODS: The cross-sectional study was conducted from February to December 2016 in the periurban low-resource locality of Bin Qasim Town in Karachi. Serum samples were screened for hepatitis B surface antigen and hepatitis C virus antibodies. Anthropometric measurements were taken and markers related to cardiovascular disease were examined. Association of the two hepatitis virus infections with cardiovascular diseasewere investigated by anaylsing the data using SPSS 16. RESULTS: There were 691 subjects. Serum triglyceride levels were significantly low in patients with hepatitis B virus (p<0.05). Those with hepatitis C virus had markedly low total cholesterol, low-density lipoprotein cholesterol, high-density lipoprotein cholesterol and triglyceride levels (p<0.05 each), whereas random blood sugar and body mass index values were significantly high (p<0.05 each. Hepatitis C virus infection was positively associated with body mass index and random blood glucose, and inversely associated with total cholesterol, high-density lipoprotein cholesterol and low-density lipoprotein cholesterol(p< 0.05 each). CONCLUSIONS: Hepatitis B virus infection showed a significant inverse association with triglyceride levels. However, hepatitis C virus infection was positively associated with body mass index and random blood sugar, and inversely associated with total cholesterol, high-density lipoprotein cholesterol and low-density lipoprotein cholesterol levels.

An Unbiased Lipid Phenotyping Approach To Study the Genetic Determinants of Lipids and Their Association with Coronary Heart Disease Risk Factors.

Direct infusion high-resolution mass spectrometry (DIHRMS) is a novel, high-throughput approach to rapidly and accurately profile hundreds of lipids in human serum without prior chromatography, facilitating in-depth lipid phenotyping for large epidemiological studies to reveal the detailed associations of individual lipids with coronary heart disease (CHD) risk factors. Intact lipid profiling by DIHRMS was performed on 5662 serum samples from healthy participants in the Pakistan Risk of Myocardial Infarction Study (PROMIS). We developed a novel semi-targeted peak-picking algorithm to detect mass-to-charge ratios in positive and negative ionization modes. We analyzed lipid partial correlations, assessed the association of lipid principal components with established CHD risk factors and genetic variants, and examined differences between lipids for a common genetic polymorphism. The DIHRMS method provided information on 360 lipids (including fatty acyls, glycerolipids, glycerophospholipids, sphingolipids, and sterol lipids), with a median coefficient of variation of 11.6% (range: 5.4-51.9). The lipids were highly correlated and exhibited a range of associations with clinical chemistry biomarkers and lifestyle factors. This platform can provide many novel insights into the effects of physiology and lifestyle on lipid metabolism, genetic determinants of lipids, and the relationship between individual lipids and CHD risk factors.

Controlled release of cephradine by biopolymers based target specific crosslinked hydrogels.

The novel silane crosslinked (TEOS) hydrogels based on eco-friendly biodegradable chitosan/guargum were prepared by blending with PEG to develop pH sensitive hydrogels (CGP) and achieved its hydrophilicity and target specificity for controlled release of drug. The crosslinker amount was varied to analyze its effect on the hydrogel properties and were characterized using FTIR, SEM, TGA, swelling studies (water, buffer and ionic solution) and in-vitro release of cephradine (CED). FTIR confirmed the presence of characteristic peaks and crosslinking between the components while SEM images showed the formation of clear micro- and macro-pores. The swelling behavior in water showed that compared to the controlled hydrogel, the crosslinked hydrogels revealed more swelling but a decrease in swelling with further increase in the amount of crosslinker was observed. The hydrogels showed low swelling at basic and neutral pH while maximum swelling was observed at acidic pH. This pH response made these hydrogels an ideal candidate for injectable controlled release. The CED was loaded on hydrogels and its release mechanism was studied in PBS, SGF and SIF which revealed that out of all hydrogels (CGP100, CGP150, CGP200 and CGP250), CGP100 has shown CED release of 85% in 130 min in PBS and 82.4% in SIF.

Co-concentration effect of silane with natural extract on biodegradable polymeric films for food packaging.

Novel biodegradable films were prepared by blending guar gum, chitosan and poly (vinyl alcohol) having mint (ME) and grapefruit peel (GE) extracts and crosslinked with nontoxic tetraethoxysilane (TEOS). The co-concentration effect of TEOS with natural extracts on the films was studied. FTIR analysis confirmed the presence of incorporated components and the developed interactions among the polymer chains. The surface morphology of the films by SEM showed the hydrophilic character due to porous network structure. The films having both ME and GE with maximum amount of crosslinker (100µL), showed maximum swelling (58g/g) and stability while the optical properties showed increased protection against UV light. This film sample showed compact network structure which enhanced the ultimate tensile strength (40.03MPa) and elongation at break (104.8%). ME/GE conferred the antioxidant properties determined by radical scavenging activity and total phenolic contents (TPC) as ME films have greater TPC compared to GE films. The soil burial test exhibited the degradation of films rapidly (6days) confirming their strong microbial activity in soil. The lower water vapour transmission rate and water vapour permeability showed better shelf life; hence, these biodegradable films are environmental friendly and have potential for food and other packaging.

Identification of new susceptibility loci for type 2 diabetes and shared etiological pathways with coronary heart disease.

To evaluate the shared genetic etiology of type 2 diabetes (T2D) and coronary heart disease (CHD), we conducted a genome-wide, multi-ancestry study of genetic variation for both diseases in up to 265,678 subjects for T2D and 260,365 subjects for CHD. We identify 16 previously unreported loci for T2D and 1 locus for CHD, including a new T2D association at a missense variant in HLA-DRB5 (odds ratio (OR) = 1.29). We show that genetically mediated increase in T2D risk also confers higher CHD risk. Joint T2D-CHD analysis identified eight variants-two of which are coding-where T2D and CHD associations appear to colocalize, including a new joint T2D-CHD association at the CCDC92 locus that also replicated for T2D. The variants associated with both outcomes implicate new pathways as well as targets of existing drugs, including icosapent ethyl and adipocyte fatty-acid-binding protein.

Development of a novel pH sensitive silane crosslinked injectable hydrogel for controlled release of neomycin sulfate.

Silane crosslinked biopolymer based novel pH-responsive hydrogels were fabricated by blending the cationic (chitosan) and anionic (alginate) polymers with poly(vinyl alcohol). Tetraethoxysilane (TEOS) was used, as a crosslinker in different amounts due to its nonhazardous nature, to study its impact on physical and chemical properties of the prepared injectable hydrogels along with the controlled release of drug. The swelling response of the prepared hydrogels was examined in different solvent media which exhibited decreased swelling ratio with increase in the amount of TEOS. All the fabricated hydrogels represented highest swelling at acidic pH while low swelling at basic and neutral pH. This specific pH sensitive behavior at pH 7 made them an appropriate candidate for the injectable controlled drug delivery in which Neomycin Sulfate (NMS) was successfully loaded on suitable hydrogel (comprising 50µL TEOS) to study its release mechanism. The results revealed that in simulated gastric fluid (SGF), hydrogel released the entire drug (NMS) in initial 30min while in simulated intestinal fluid (SIF), NMS was released in a controlled way up to 83% in 80min. These results endorsed that the hydrogels could be practiced as a smart intelligent material for injectable controlled drug delivery as well as for other biomedical applications at physiological pH.